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Toxicity to aquatic algae and cyanobacteria

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Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Justification for type of information:
See Section 13 RAAF reports for the justification of read-across.
Reason / purpose:
read-across source
Remarks:
6.1.5.027
Reason / purpose:
read-across source
Remarks:
6.1.5.110
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 684 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Remarks:
dimethylsilanol
Basis for effect:
growth rate
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
44 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Remarks:
dimethylsilanol
Basis for effect:
growth rate
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
135 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Remarks:
dimethylsilanol
Basis for effect:
cell number
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
683 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Remarks:
dimethylsilanol
Basis for effect:
cell number
Remarks on result:
other: 641-727
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
70 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Remarks:
dimethylsilanol
Basis for effect:
biomass
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
625 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Remarks:
dimethylsilanol
Basis for effect:
biomass
Remarks on result:
other: 555-702
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
522 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Remarks:
dimethylsilanol
Basis for effect:
growth rate
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
> 1 053 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Remarks:
dimethylsilanol
Basis for effect:
growth rate
Remarks on result:
other: not calculable
Key result
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
> 118 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Remarks:
DMSD
Basis for effect:
other: growth rate and cell yield
Key result
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
>= 118 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Remarks:
DMSD
Basis for effect:
other: yield and growth rate
Validity criteria fulfilled:
yes
Conclusions:
A 72-hour EC50 value of >1053 mg/L and NOEC of 70 mg/L have been determined for the effects of the test on growth rate of Selenastrum capricornutum (new name: Pseudokirchneriella subcapitata) based on intitial measured concentrations of the test substance. When expressed relative to mean measured concentrations the equivalent values are >750 mg/L and 50 mg/L.
Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
2004-03-11 to 2004-03-15
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
Qualifier:
according to
Guideline:
EU Method C.3 (Algal Inhibition test)
Qualifier:
according to
Guideline:
EPA OPPTS 850.5400 (Algal Toxicity, Tiers I and II)
GLP compliance:
yes
Analytical monitoring:
yes
Details on sampling:
- Concentrations: Negative Control, 31, 63, 125, 250, 500 and 1000 mg/L

- Sampling method: Test concentrations were measured in each treatment at test initiation and at test termination.



Vehicle:
no
Details on test solutions:
PREPARATION AND APPLICATION OF TEST SOLUTION

- Method: Direct addition of test article to freshwater algal medium

- Controls: Freshwater Algal Medium (OECD 201)
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM

- Source (laboratory, culture collection): Original algal cultures were obtained from the University of Toronto Culture Collection, and had been maintained in culture medium at Wildlife International, Ltd., Easton, Maryland. Algal cells used in this test were obtained from Wildlife International, Ltd. cultures that had been actively growing in culture medium for at least two weeks prior to test initiation. The culture was last transferred to fresh medium three days prior to test initiation.

- Culture medium: The algal cells were cultured and tested in freshwater algal medium. Stock nutrient solutions were prepared by adding reagent-grade chemicals to purified Wildlife International, Ltd. well water. The test medium then was prepared by adding appropriate volumes of the stock nutrient solutions to purified well water (NANOpure® water). The pH of the medium was adjusted to 7.9 using 10% HCl and the medium was sterilized by filtration (0.22 μm) prior to use.

- Initial density: 10,000 cells/mL
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
96 h
Post exposure observation period:
None
Test temperature:
23.6 to 24.6 ºC



pH:
7.9 at test initiation

8.3 to 9.9 at test termination
Dissolved oxygen:
Not applicable
Salinity:
Not applicable
Nominal and measured concentrations:
Nominal concentrations: Negative Control, 31, 63, 125, 250, 500 and 1000 mg/L.

Measured concentrations:
Test concentrations declined to less than 70% of nominal at test termination. Consequently, the results of the study were based on Day 0 measured concentrations.
Details on test conditions:
TEST SYSTEM

- Test vessel: 250-mL Erlenmeyer flasks plugged with cotton stoppers containing 100 mL of test solution.

- Aeration: none

- Renewal rate of test solution (frequency/flow rate): static test

- Initial cells density: 10000 cells/mL

- No. of vessels per concentration (replicates): 3

- No. of vessels per control (replicates): 6


GROWTH MEDIUM

- Standard medium used: yes


TEST MEDIUM / WATER PARAMETERS

- Source/preparation of dilution water: Freshwater algal growth medium (OECD 201) with NANOpure water (pH: 7.9)

- Culture medium different from test medium: no

- Intervals of water quality measurement: Start and end of test


OTHER TEST CONDITIONS

- Sterile test conditions: yes

- Adjustment of pH: no

- Photoperiod: Continuous

- Light intensity and quality: 3900 to 4700 lux, continuous cool-white fluorescent lighting


EFFECT PARAMETERS MEASURED: Cell densities measured at start of test and then at 24 hour intervals. Cell densities were used to determine effects on average-specific growth rate and biomass.


TEST CONCENTRATIONS

- Spacing factor for test concentrations: 2
Reference substance (positive control):
no
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
70 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
cell number
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
555 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
cell number
Remarks on result:
other: 141-612
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
135 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
cell number
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
683 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
cell number
Remarks on result:
other: 641-727
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
70 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
biomass
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
566 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks on result:
other: 409-618
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
70 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
biomass
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
625 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
biomass
Remarks on result:
other: 555-702
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
70 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 1 053 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: not calculable
Duration:
96 h
Dose descriptor:
NOEC
Effect conc.:
522 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
96 h
Dose descriptor:
EC50
Effect conc.:
> 1 053 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: not calculable
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
50 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 750 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Remarks on result:
other: not calculable
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 684 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
44 mg/L
Nominal / measured:
meas. (geom. mean)
Conc. based on:
test mat.
Basis for effect:
growth rate
Details on results:
Was control response satisfactory: Yes

Observations: There were no noticeable changes in cell morphology in any of the tested concentrations in comparison to the control.



Reported statistics and error estimates:
Statistical methods: Non-linear regression, linear interpolation and Dunnett's test

Table 1. Test results (cell densities in cells/mL)

Day 0 Measured  Concentration (mg/L)
          0 33     70     135    257    522    1053
0-Hour  10000  10000 10000 10000  10000  10000  10000
24-Hour  72000 75000 67000 67000 60000 51000 22000
48-Hour  396000 389000   345000 322000  263000  193000  44000
72-Hour  1780000 1700000 1680000 1090000 882000  995000  172000
96-Hour  4660000 4620000 4690000 4740000 3810000 3500000 570000

Table 2. Percent biomass/growth rate inhibition per concentration


         Day 0 Measured Concentration (mg/L)
                 33    70    135   257   522   1053
72-Hour Cell Density  4.6   5.8   39    51    44    90
96-Hour Cell Density  0.74 -0.70 -1.8   18    25    88
72-Hour Biomass       3.4   8.0   32    45    46    90
96-Hour Biomass       2.3   3.1   16    32    35    89
72-Hour Growth Rate   0.94  1.2   9.5   14    11    45
96-Hour Growth Rate   0.12 -0.11 -0.30  3.3   4.6   34

Validity criteria fulfilled:
yes
Conclusions:
A 72-hour EC50 value of >1053 mg/L and NOEC of 70 mg/L have been determined for the effects of the test on growth rate of Selenastrum capricornutum (new name: Pseudokirchnerella subcapitata) based on initial measured concentrations of the test substance. When expressed relative to geometric mean measured concentrations the equivalent values are >684 mg/L and 44 mg/L.
Endpoint:
toxicity to aquatic algae and cyanobacteria
Type of information:
experimental study
Adequacy of study:
weight of evidence
Study period:
2008-08-05 to 2008-08-14
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 201 (Alga, Growth Inhibition Test)
GLP compliance:
yes (incl. certificate)
Analytical monitoring:
yes
Details on sampling:
- Sampling method: Samples of each test medium were taken at the start and end of the test. They received no further treatment prior to analysis.
Vehicle:
no
Details on test solutions:
Nominal test concentrations were 7.5, 15, 30, 60 and 120 mg/L. A primary stock was prepared by adding 0.12 g of the test article directly to the algal assay medium (AAM) in a 1-L volumetric flask. The flask was brought to volume with AAM and inverted to mix. Four secondary stock solutions were prepared at concentrations of 60, 30, 15 and 7.5 mg/L by 50% serial dilutions of the primary stock. Each stock was inverted to mix. All stock solutions were clear and colorless. Approximately 100 mL of test solution was added to each of the three replicates and the media blank for each treatment group. Remaining test solution in the volumetric flasks was used for initial water quality measurements.
Test organisms (species):
Pseudokirchneriella subcapitata (previous names: Raphidocelis subcapitata, Selenastrum capricornutum)
Details on test organisms:
TEST ORGANISM

- Source (laboratory, culture collection): Test organisms were obtained from in-house cultures. The source of the original brood stock was strain UTEX 1648, from the University of Texas at Austin, The Culture Collection of Algae, Botany Department, Austin, Texas, in March 1998. The identity of the species was verified by the supplier.

- Method of cultivation: Algal cultures were maintained at 24+/-2ºC on a rotary shaker set at approximately 100 rpm, under continuous cool white fluorescent lighting at a range of 400 ± 100 foot candles (4300+/-1075 lux). The cultures were periodically transferred axenically to new AAM.

- Culture medium: Algal Assay Medium was prepared based on ASTM guideline E 1218-04. The final concentration of each nutrient in the medium is presented in Appendix A. All stocks used to prepare the medium were prepared with sterile distilled water and were filter sterilized (0.22 μm). Medium was prepared with commercially available distilled water that is analyzed annually for contaminants. No contaminants have been shown to be present which might adversely impact the viability of the cultures or study
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
72 h
Hardness:
Not reported
Test temperature:
23.8 to 24.0ºC
pH:
pH at test initiation ranged from 7.4 to 7.5. At test termination, pH in the flasks that contained algae ranged from 8.0 to 9.0 and pH of media blanks ranged from 7.6 to 7.8
Dissolved oxygen:
Not reported
Salinity:
Not applicable
Nominal and measured concentrations:
Nominal test concentrations: 0 (Control), 7.5, 15, 30, 60 and 120 mg/L

Mean measured test concentrations in treated media: 8, 18, 28, 57 and 118 mg/L; which is 106, 122, 93, 96 and 99% of nominal.

The results are expressed relative to mean measured concentrations.
Details on test conditions:
TEST SYSTEM

- Test Apparatus: Test vessels were sterile 250-mL Erlenmeyer flasks covered with a sponge closure containing approximately 100 mL of test solution. Test vessels were labeled with the study number, concentration, and replicate.

- Initial cell density: 10000 cells/ml

- Test Conditions: Test vessels were placed in a temperature controlled incubator and positioned on rotary shakers that were set at approximately 100 rpm. In addition, the incubator was set at a temperature of 24+/-2ºC and under continuous cool white fluorescent lighting at a range of 446 – 620 foot-candles.

Temperature was measured in a beaker of water adjacent to the shakers in the incubator and light readings were recorded at five indiscriminate areas on the shaker tables daily during the test. Readings of pH were completed on bulk preparations of each treatment group at test initiation. To maintain axenic conditions, pH readings for each test vessel were measured only at 72 hours.

- Observations: Samples for cell counts were collected on Day 0 for the original culture and test inoculums. Samples for cell counts were also collected from each test vessel at 24, 48 and 72 hours (within 1 hour). Samples were diluted with Isoton® II diluent solution and counted immediately using a Coulter® Counter. Dilution volumes were recorded in the raw data. Dilution samples of the medium and diluent blanks were counted at 24, 48, and 72 hours to account for any interference of particles. At test termination, samples were pooled by treatment and visually inspected for atypical cell morphology. Visual examination showed no concentration-dependent changes in shape, color or size.

- Light intensity: 446 to 620 foot-candles
Reference substance (positive control):
no
Key result
Duration:
72 h
Dose descriptor:
EC50
Effect conc.:
> 118 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
other: yield and growth rate
Key result
Duration:
72 h
Dose descriptor:
NOEC
Effect conc.:
>= 118 mg/L
Nominal / measured:
meas. (arithm. mean)
Conc. based on:
test mat.
Basis for effect:
other: yield and growth rate
Details on results:
- Exponential growth in the control (for algal test): yes, Control cell density for this study increased 122 fold by 72 hours.
Reported statistics and error estimates:
ECx values for yield and growth rate at 72 hours were calculated using linear interpolation or estimated by visual examination of the data.

Yield (biomass) and growth rate data were analyzed for normality and homogeneity of variance using the Shapiro-Wilk’s test and Bartlett’s test, respectively. The no observed effect concentration (NOEC) values were determined for growth rate and yield at 72 hours using Dunnett’s test. All statistical analyses were performed using TOXSTAT Version 3.5 software

Table 1. Measurement of Test Concentrations

Nominal

Concentration

(mg/L)

Day 0 Measured Concentration

 (mg/L)

Day 3 Measured Concentration (mg/L)

Mean

Measured

Concentration

(mg/L)

Percent of Nominal

Control

 

ND

ND

--

--

7.5

 

5.9

10

8.0

106

15

 

18

19

18

122

30

 

25

31

28

93

60

55

60

57

96

120

114

122

118

99

ND = Not Detected

 

Table 2. Mean Cell Densities (cells/mL)

Mean Measured Concentration

(mg/L)

 

 

0-hour

24-hour

48-hour

72-hour

Negative Control

~10000

43433

240424

1222333

8

~10000

41551

246551

1276867

18

~10000

37384

232924

1216000

28

~10000

38551

217216

1129333

57

~10000

42022

228560

1090778

118

~10000

35773

201429

990800

Algal cell growth in the control was exponential during the 72-h exposure period. Cell densities were used to calculate the average specific growth rates and yield for each test concentration. 

Validity criteria fulfilled:
yes
Conclusions:
A 72-hour EC50 value of >118 mg/L and a NOEC of ≥118 mg/L have been determined for the effects of the test substance on yield and growth rate of Pseudokirchneriella subcapitata.

Description of key information

Toxicity to algae: 72 hr EC50 >684 mg/l (highest concentration tested) and NOEC 44 mg/l based on geometric mean measured concentrations, in accordance with OECD guideline 201, read-across from an analogous/structurally related substance trimethylsilanol (CAS 1066-40-6).

Key value for chemical safety assessment

EC10 or NOEC for freshwater algae:
44 mg/L

Additional information

There are no test data for the registered substance. However data are available from tests carried out with trimethylsilanol (CAS 1066-40-6), a structurally analogous substance to the intermediate hydrolysis product dimethylsilanol. Data are also available with the ultimate hydrolysis product dimethylsilanediol (CAS 1066-42-8).

Data with trimethylsilanol:

  • A 72-hour EC50 value of >1053 mg/l and NOEC of 70 mg/l have been determined for the effects of trimethylsilanol on growth rate of Pseudokirchneriella subcapitata based on initial measured concentrations of the test substance. When expressed relative to geometric mean measured concentrations the equivalent values are >684 mg/l and 44 mg/l.
  • The study is supported by another study in which a 72-hour algal EC50 value of >1000 mg/l and NOEC of 316 mg/l (Bayer 1992).

For trimethylsilanol, a solution at 100 mg/l is predicted to contain >99.9% monomer. At loadings above about 500 - 1000 mg/l the concentration of the dimer is predicted to exceed its solubility, resulting in formation of a separate phase. During the algae test with trimethylsilanol, less than 20% growth inhibition was observed up to the 522 mg/l loading level and the EC50 is considered to be above the highest concentration tested. In addition, the dimer that could be formed due to the condensation of two trimethylsilanol molecules is hexamethyldisiloxane (HMDS, CAS 107-46-0). HMDS is classified as Aquatic Acute 1 H400 and Aquatic Chronic 2 H411[1] and therefore its presence may impact the toxicity results. However, the estimated concentration of HMDS is 0.04, 0.2, 0.3, 0.6 and 0.9 mg/l at 100, 200, 300, 400 and 500 mg/l of TMS respectively, and there was no reporting of undissolved test material or droplets being formed, which would otherwise clearly indicate the formation of HMDS. Furthermore, the log Kow of trimethylsilanol (1.62) is such that effects at relatively high loadings may be observed. Therefore, any condensation reactions that may have occurred did not significantly impact the results of the study and it is concluded that the study is considered to be reliable for the purpose of this assessment. This conclusion is supported by the analogous EC50 results obtained in the two algal studies with TMS.

Data with dimethylsilanediol:

-       A 72-hour EC50 value of >118 mg/l and a NOEC of ≥118 mg/l have been determined for the effects of dimethylsilanediol on growth rate of Pseudokirchneriella subcapitata based on mean measured concentrations.

The registered substance also releases hydrogen during its final hydrolysis. Hydrogen is not thought to contribute to the toxicity of the substance and is discussed further in the ecotoxicological information overview endpoint summary (additional information).

Details on how the PNEC and the risk characterisation ratio have been derived can be found in IUCLID Section 6.0 and Chapters 9 and 10 of the Chemical Safety Report, respectively.   


[1] https://echa.europa.eu/information-on-chemicals/cl-inventory-database/-/discli/notification-details/32082/948794